Our work focuses primarily on a regulatory system in the yeast Saccharomyces, known as general amino acid control, which governs the transcription of a large number of amino acid biosynthetic genes in response to amino acid availability. Our recent results extend our previous finding that the proximal trans-acting positive effector in this system (GCN4) is itself regulated and this regulation occurs at the translational level. Analysis of a GCN4-lacZ gene fusion indicates that all other known general control trans-acting factors exert their effects indirectly as translational effectors of GCN4. This was shown directly by the fact that their effects on GCN4 expression are independent of the GCN4 promoter. GCN4 mRNA contains a cis-acting negative element which mediates the translational control of GCN4. This site encompasses 4 short open-reading-frames (ORFs). We have generated point mutations in the AUG codons of these ORFs and preliminary results suggest that these sequences are the primary regulatory elements in the GCN4 mRNA leader. Interestingly, the ORFs are not equivalent in their effects and the first ORF seems to be critical for establishing the regulatory effects of the trans-acting factors on GCN4 translation. We have also isolated four new complementation groups of mutations which inactivate or bypass the GCN4 short ORFs. One of the groups is closely linked to GCN4 and may represent cis-acting constitutive alleles of GCN4. Finally, we are attempting to produce antisera specific for the protein products of GCN4 and its positive regulator encoded by GCN3. These antisera will be used to determine the cellular localization and for purification of these proteins. We are also involved in collaborative work on the expression of the yeast homologues of the RAS cellular oncogenes and the RNase H gene of yeast.